IGF1-mediated human embryonic stem cell self-renewal recapitulates the embryonic niche

Sissy E Wamaitha,Julian Downward,Miriam Molina-Arcas,Liani G Devito,James M A Turner,Kay Elder,Rebecca A Lea,Sugako Ogushi,Leila Christie,Shantha K Mahadevaiah,Lyn Healy,Phil Snell,Gregorio Alanis-Lobato,Claudia Gerri,Kathy K Niakan,Afshan Mccarthy,Katarzyna J Grybel

doi:10.1038/s41467-020-14629-x

Abstract

Our understanding of the signalling pathways regulating early human development is limited, despite their fundamental biological importance. Here, we mine transcriptomics datasets to investigate signalling in the human embryo and identify expression for the insulin and insulin growth factor 1 (IGF1) receptors, along with IGF1 ligand. Consequently, we generate a minimal chemically-defined culture medium in which IGF1 together with Activin maintain self-renewal in the absence of fibroblast growth factor (FGF) signalling. Under these conditions, we derive several pluripotent stem cell lines that express pluripotency-associated genes, retain high viability and a normal karyotype, and can be genetically modified or differentiated into multiple cell lineages. We also identify active phosphoinositide 3-kinase (PI3K)/AKT/mTOR signalling in early human embryos, and in both primed and naïve pluripotent culture conditions. This demonstrates that signalling insights from human blastocysts can be used to define culture conditions that more closely recapitulate the embryonic niche.

Highlights

Our understanding of the signalling pathways regulating early human development is limited, despite their fundamental biological importance
We identify active IGF signalling via phosphoinositide 3-kinase (PI3K)/mTOR in human preimplantation embryos, and in both conventional and naïve human embryonic stem cells (hESCs) culture conditions, and determine that inhibiting fibroblast growth factor (FGF) receptors does not adversely affect hESC maintenance
We previously showed that several TGFβ/NODAL signallingassociated genes were enriched in the EPI, and that intact TGFβ/Nodal signalling is required for pluripotency gene expression in the human embryo[10], as had been described in hESCs13

Summary

Introduction

Our understanding of the signalling pathways regulating early human development is limited, despite their fundamental biological importance. We generate a minimal chemically-defined culture medium in which IGF1 together with Activin maintain self-renewal in the absence of fibroblast growth factor (FGF) signalling. Under these conditions, we derive several pluripotent stem cell lines that express pluripotency-associated genes, retain high viability and a normal karyotype, and can be genetically modified or differentiated into multiple cell lineages. TGFβ/Nodal signalling is active during early human development and has been implicated in maintaining NANOG expression in the human EPI10, as well as regulating pluripotency gene expression in human embryonic stem cells (hESCs)[11,12,13,14,15]. FGF is included in the majority of hESC culture medium, either by the addition of exogenous ligand, or by co-culture with mitotically inactivated fibroblast (MEF) layers, use of MEF-conditioned medium, or growth on a Matrigel matrix[22,23,24,25,26,27]

Methods

Results

Conclusion